Ribosome rescue factor PELOTA modulates translation start site choice for C/EBPα protein isoforms

Abstract

Translation initiation at alternative start sites can dynamically control the synthesis of two or more functionally distinct protein isoforms from a single mRNA. Alternate isoforms of the developmental transcription factor CCAAT/enhancer-binding protein α (C/EBPα) produced from different start sites exert opposing effects during myeloid cell development. This choice between alternative start sites depends on sequence features of the CEBPA transcript, including a regulatory uORF, but the molecular basis is not fully understood. Here, we identify the factors that affect C/ EBPα isoform choice using a sensitive and quantitative two-color fluorescent reporter coupled with CRISPRi screening. Our screen uncovered a role of the ribosome rescue factor PELOTA (PELO) in promoting the expression of the longer C/EBPα isoform by di-rectly removing inhibitory unrecycled ribosomes and through indirect effects mediated by the mechanistic target of rapamycin kinase. Our work uncovers further links between ribosome recycling and translation reinitiation that regulate a key transcription factor, with implications for normal hematopoiesis and leukemogenesis.